Anti-MT drugs continue to receive great interest for their development as potential clinically useful antitumor agents. Several new natural products with taxol-like activity towards tubulin and microtubules have been identified recently. Of these, the epothiones are the most promising and exciting. This group of compounds can be isolated from bacterial source, making large scale production feasible. Epothilone A and B have been shown to be equipotent to taxol in stimulating tubulin assembly and competitively inhibiting [3H]taxol binding to microtubules. The epothilones are also similar to taxol in their anti-cell proliferative activity. However, these compounds have much greater activity toward cells that are resistant to taxol due to the over-expression of the P-glycoprotein multidrug transporter or to the expression of an altered Beta-tubulin. Thus, these compounds have the potential of overcoming two different mechanisms of cell resistance to taxol. Very little is known yet about the interaction of drugs with microtubules. Such information would be useful for designing new and better derivatives. Information obtained on the binding site will also be extremely important for future work on the site directed mutagenesis of the tubulin protein. A variety of approaches can be taken to obtain this information, including those which we are proposing, identification of peptides in the binding site through photo affinity labeling and the use of fluorescence spectroscopy to study detail of the interaction. The results of the tubulin binding studies would suggest that the binding sites for taxol and epothilone are identical. However, the fact that the epothilones are effective cytotoxic agents in cells that are resistant to taxol because of the expression of an altered Beta-tubulin, indicates that the sites may be overlapping rather than identical. It is the plan to prepare epothilone affinity labels through emisynthesis and total synthesis and evaluate them for tubulin assembly activity and cytotoxicity. Fluorescent epothilones will be used to study the interaction with microtubules and tubulin and the epothilone photo affinity will identify the ligand binding sites in the receptor.